The long term goal of this project is to elucidate the mechanism of receptor regulation of adrenal hormone secretion in isolated and cultured adrenal chromaffin cells. The chromaffin cell has long been considered a paradigm of stimulus-secretion coupling and much of our general knowledge of cellular secretory mechanisms has been derived from studies of the perfused adrenal gland. The recent availability of isolated and cultured chromaffin cells promises new insights into secretroy mechanisms in general and catecholamine release in particular. The roles of several important receptor mediated membrane responses in regulating secretion will be investigated including: 1) activation of Na, Ca, and cholinergic receptor ion channels, 2) membrane action potential spiking, 3) calcium translocation and 4) alterations in levels of cyclic AMP and cyclic GMP. Chromaffin cells will be isolated by sequential collagenase digestion of bovine adrenal medulla and maintained in culture on collagen coated dishes in Dulbecco's Modified Eagles Medium supplemented with 10% fetal calf serum. Catecholamine secretion will be measured by a (3H) norepenephrine assay and independently by the standard THI fluorescence assay. Membrane electric potential will be monitored by glass microelectrodes and independently using potential sensitive optical probes. Membrane calcium translocation will be monitored by 45Ca tracer techniques and by fluorescent probes. A variety of pharmacological agents will be used to influence ion channels, membrane potential, cyclic nucleological levels, and cholinergic receptors and their corresponding effects upon catecholamine secretion will be determined. An improved understanding of the mechanism and pharmacology of cellular secretion should help in the management of certain endocrine and exocrine diseases having defective secretion, notably cystic fibrosis, hyperinsulinism, acromegaly, and the adrenal medullary tumor, pheochromocytoma. There is also the direct relevance of adrenaline release to pulmonary and cardiovascular function and hypertention.